Mammalian chaperonin 10 (Cpn10), also known as heat shock protein 10 (Hsp10) and early pregnancy factor (EPF), is typically characterised as a mitochondrial ‘molecular chaperone’ protein involved in protein folding together with chaperonin 60 (Cpn60), also known as heat shock protein 60 (Hsp60). Cpn10 and Cpn60 are homologues of the bacterial proteins GroES and GroEL respectively. GroES and Cpn10 each oligomerise into seven member rings that bind as a lid onto a barrel-like structure comprising fourteen GroEL or seven Cpn60 molecules respectively, which tether denatured proteins to the complex (Bukau and Horwich, 1998, Cell 92:351-366; Hartl and is Hayer-Hartl, 2002, Science 295:1852-1858).
Cpn10 proteins are highly conserved across species. Human Cpn10 is 100% identical to bovine, canine, ovine and porcine Cpn10 and differs from rat Cpn10 at only a single amino acid position. Human Cpn10 shares 38% sequence identity (60% similarity) with GroES from Escherichia coli. Cpn10/GroES proteins are dome shaped heptameric rings wherein each monomer is comprised of essentially three different structural regions, a core anti-parallel β-barrel region flanked by a “roof” β-hairpin loop region and a “mobile loop” region. The anti-parallel β-barrel region of each monomer forms the core of a dome and when assembled in the heptamer the β-hairpin loops of each monomer form the roof of the dome. In each monomer, the mobile loop region is at the opposite end of the β-barrel to the roof loops. A section of the anti-parallel β-barrel region forms an inward facing lower rim region of the cavity. This lower rim region contains a number of phylogenetically conserved amino acids including a Tyrosine at position 75 (Y75).
In addition to its intracellular role as a molecular chaperone, Cpn10 is also frequently found at the cell surface (see Belles et al., 1999, Infect lmmun 67:4191-4200) and in the extracellular fluid (see Shin et al., 2003, J Biol Chem 278:7607-7616) and is increasingly being recognised as a regulator of the immune response with potential in the treatment of inflammatory disorders. Accordingly, the efficacy and safety of Cpn10 has recently been established in the treatment of human patients with rheumatoid arthritis (Vanags et al. Lancet 2006, 368: 855-863) and psoriasis (Williams et al. Arch. Dermatol. 2008, 144: 683-685).
However the sites within the Cpn10 molecule responsible for mediating this immunomodulatory activity have remained elusive. The present invention relates to the discovery that modification of Cpn10 affects the immunomodulatory activity of Cpn10, in particular its role in binding ligands of pattern recognition receptors (PRRs) such as Toll-like Receptors (TLR), Nucleotide-binding domain LRR-containing family (NLR), RIG-I-like receptors (RLR), DNA-dependent activators of IRF (DAI), C-type Lectin receptors (CLR) or a member of the IFI20X/IFI16 family (e.g. Ifi16, Aim2, MNDA and IFIX).